Uptake of osteoblast-derived extracellular vesicles promotes the differentiation of osteoclasts in the zebrafish scale

Differentiation of osteoclasts (OCs) from hematopoietic cells requires cellular interaction with osteoblasts (OBs). Due to the difficulty of live-imaging in the bone, however, the cellular and molecular mechanisms underlying intercellular communication involved in OC differentiation are still elusive. Here, we develop a fracture healing model using the scale of trap:GFP; osterix:mCherry transgenic zebrafish to visualize the interaction between OCs and OBs. Transplantation assays followed by flow cytometric analysis reveal that most trap:GFP(high) OCs in the fractured scale are detected in the osterix:mCherry(+) fraction because of uptake of OB-derived extracellular vesicles (EVs). In vivo live-imaging shows that immature OCs actively interact with osterix:mCherry(+) OBs and engulf EVs prior to convergence at the fracture site. In vitro cell culture assays show that OB-derived EVs promote OC differentiation via Rankl signaling. Collectively, these data suggest that EV-mediated intercellular communication with OBs plays an important role in the differentiation of OCs in bone tissue.